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          细胞工程大纲
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        <h1 id="细胞工程"><a href="#细胞工程" class="headerlink" title="细胞工程"></a>细胞工程</h1><h2 id="前言"><a href="#前言" class="headerlink" title="前言"></a>前言</h2><h3 id="原理和方法"><a href="#原理和方法" class="headerlink" title="原理和方法"></a>原理和方法</h3><ul>
<li>细胞生物学</li>
<li>分子生物学</li>
</ul>
<h3 id="操作水平"><a href="#操作水平" class="headerlink" title="操作水平"></a>操作水平</h3><ul>
<li>细胞水平</li>
<li>细胞器水平</li>
</ul>
<h3 id="目的"><a href="#目的" class="headerlink" title="目的"></a>目的</h3><ul>
<li>改变细胞内的遗传物质</li>
<li>获得细胞产品</li>
</ul>
<h3 id="两大领域"><a href="#两大领域" class="headerlink" title="两大领域"></a>两大领域</h3><ul>
<li>植物细胞工程</li>
<li>动物细胞工程</li>
</ul>
<h2 id="植物细胞工程"><a href="#植物细胞工程" class="headerlink" title="植物细胞工程"></a>植物细胞工程</h2><h3 id="理论基础"><a href="#理论基础" class="headerlink" title="理论基础"></a>理论基础</h3><ul>
<li>细胞全能性</li>
</ul>
<h3 id="相关概念"><a href="#相关概念" class="headerlink" title="相关概念"></a>相关概念</h3><ul>
<li><p>外植体</p>
<ul>
<li>用于组培的离体的植物器官、组织、细胞</li>
</ul>
</li>
<li><p>脱分化</p>
<ul>
<li>让已经分化的细胞，经过诱导后，失去其特有的结构和功能而转变成未分化细胞的过程。</li>
</ul>
</li>
<li><p>愈伤组织</p>
<ul>
<li>排列疏松，高度液泡化，呈无定形状态的薄壁细胞团</li>
</ul>
</li>
<li><p>丛芽（胚状体）</p>
<ul>
<li>由愈伤组织再分化得到</li>
</ul>
</li>
<li><p>全能性</p>
<ul>
<li><p>概念</p>
<ul>
<li>具有某种生物全部遗传信息的任何一个细胞，都具有发育成完整生物体的潜能，也就是说，每个生物细胞都具有全能性的特点。</li>
</ul>
</li>
<li><p>表达条件</p>
<ul>
<li>离体细胞、组织、器官</li>
<li>无菌条件（避免微生物的污染）</li>
<li><p>适宜的营养（培养基）</p>
<ul>
<li>水</li>
<li>无机盐</li>
<li><p>蔗糖</p>
<ul>
<li>提供营养和调节渗透压</li>
</ul>
</li>
<li><p>……</p>
</li>
</ul>
</li>
<li><p>适宜的温度、pH、光照调节（分时段）</p>
</li>
<li><p>适宜的激素诱导</p>
<ul>
<li>生长素</li>
<li>细胞分裂素</li>
<li><p>PS：相对浓度效果</p>
<ul>
<li><p>生长素 &gt; 细胞分类素</p>
<ul>
<li>诱导脱分化</li>
<li>诱导根原基的形成</li>
</ul>
</li>
<li><p>细胞分裂素 &gt; 生长素</p>
<ul>
<li>诱导再分化</li>
<li>诱导芽原基的形成</li>
</ul>
</li>
</ul>
</li>
</ul>
</li>
</ul>
</li>
<li><p>不具有全能性的细胞</p>
<ul>
<li>没有细胞核的细胞（成熟红细胞、筛管细胞）</li>
<li>死细胞（导管细胞）</li>
</ul>
</li>
<li><p>PS：在个体发育过程中，细胞并不会表现出全能性，而是分化成各种组织和器官。</p>
</li>
</ul>
</li>
</ul>
<h3 id="基本技术"><a href="#基本技术" class="headerlink" title="基本技术"></a>基本技术</h3><ul>
<li><p>植物组织培养技术</p>
<ul>
<li>概念：在无菌和人工控制条件下，将离体的植物器官、组织、细胞，培养在人工配制的培养基上，给予适宜的培养条件，诱导其产生愈伤组织、丛芽（胚状体），最终形成完整的植株。</li>
<li><p>流程</p>
<ul>
<li><p>1.外植体脱分化得到愈伤组织</p>
<ul>
<li>胡萝卜选分化程度较低的形成层</li>
<li>愈伤组织在恒温避光条件下培养（光照条件下易分化出维管组织）</li>
</ul>
</li>
<li><p>2.愈伤组织再分化得到胚状体</p>
<ul>
<li>要有光照</li>
</ul>
</li>
<li><p>3.胚状体发育成完整植株</p>
<ul>
<li>移栽到大田</li>
</ul>
</li>
</ul>
</li>
<li><p>培养基的组成（固体）</p>
<ul>
<li>无机营养成分</li>
<li>有机营养成分</li>
<li>激素</li>
<li>琼脂</li>
</ul>
</li>
</ul>
</li>
<li><p>植物体细胞杂交技术</p>
<ul>
<li><p>概念</p>
<ul>
<li>将不同种的植物体细胞，在一定条件下融合成杂种细胞，并把杂种细胞培育成新的植物体的技术</li>
</ul>
</li>
<li><p>流程</p>
<ul>
<li><p>1.去细胞壁得到原生质体</p>
<ul>
<li>纤维素酶和果胶酶</li>
</ul>
</li>
<li><p>2.人工诱导融合</p>
<ul>
<li>体现细胞膜的流动性</li>
<li><p>方法</p>
<ul>
<li>物理法（离心、振动、电激）</li>
<li>化学法（聚乙二醇PEG）</li>
</ul>
</li>
</ul>
</li>
<li><p>3.杂种细胞再生出细胞壁</p>
<ul>
<li>真正完成融合的标志</li>
<li>需要高尔基体</li>
</ul>
</li>
<li><p>4.进行植物组织培养</p>
</li>
</ul>
</li>
<li><p>优势</p>
<ul>
<li>克服不同生物远缘杂交的障碍</li>
</ul>
</li>
</ul>
</li>
</ul>
<h3 id="实际应用"><a href="#实际应用" class="headerlink" title="实际应用"></a>实际应用</h3><ul>
<li><p>植物繁殖的新途径</p>
<ul>
<li><p>微型繁殖技术（快速繁殖技术）</p>
<ul>
<li><p>本质</p>
<ul>
<li>组织培养</li>
</ul>
</li>
<li><p>优势</p>
<ul>
<li>保持优良品种的遗传特性</li>
<li>高效快速地实现种苗的大量繁殖</li>
</ul>
</li>
</ul>
</li>
<li><p>作物脱毒</p>
<ul>
<li>无性繁殖的作物感染的病毒容易传播给后代。<br>e.g.马铃薯、草莓</li>
<li>分生区（茎尖、芽尖、根尖）附近病毒极少，取之进行组织培养获得脱毒苗</li>
<li>目前采用茎尖培养技术来脱除病毒</li>
</ul>
</li>
<li><p>人工种子</p>
<ul>
<li><p>结构</p>
<ul>
<li>人工种皮</li>
<li>人工胚乳</li>
<li><p>胚状体</p>
<ul>
<li>不定芽</li>
<li>顶芽</li>
<li>腋芽</li>
</ul>
</li>
</ul>
</li>
<li><p>优点</p>
<ul>
<li>后代无性状分离，保持优良特性</li>
<li>不受气候、地域、季节限制</li>
<li>时间短、产地少、储存方便</li>
</ul>
</li>
<li><p>人工种皮中应有的有效成分</p>
<ul>
<li>适量养分、无机盐、有机碳源</li>
<li>农药、抗生素、有益菌、植物生长调节剂</li>
</ul>
</li>
<li><p>PS：无性繁殖，不一定可育</p>
</li>
</ul>
</li>
</ul>
</li>
<li><p>作物新品种的培育</p>
<ul>
<li><p>单倍体育种</p>
<ul>
<li><p>原理</p>
<ul>
<li>细胞全能性</li>
<li>染色体数目变异</li>
</ul>
</li>
<li><p>优点</p>
<ul>
<li>可得到稳定遗传的优良品种</li>
<li>极大地缩短了育种年限</li>
</ul>
</li>
</ul>
</li>
<li><p>突变体的利用</p>
<ul>
<li><p>原理</p>
<ul>
<li>基因突变</li>
<li>植物细胞全能性</li>
</ul>
</li>
<li><p>方法</p>
<ul>
<li>对愈伤组织进行化学或物理的诱变处理</li>
</ul>
</li>
<li><p>诱变方法</p>
<ul>
<li><p>化学诱变剂</p>
<ul>
<li>甲基硫酸乙酯（EMS）</li>
<li>叠氮化钠（SA）</li>
</ul>
</li>
<li><p>物理诱变</p>
<ul>
<li><p>电离辐射</p>
<ul>
<li>γ射线</li>
<li>β射线</li>
<li>中子</li>
</ul>
</li>
</ul>
</li>
</ul>
</li>
</ul>
</li>
</ul>
</li>
<li><p>细胞产物的工厂化生产</p>
<ul>
<li><p>优点</p>
<ul>
<li>没有时间、地点、环境的影响</li>
<li>克服了植物生长缓慢的缺点</li>
<li>提取方便</li>
</ul>
</li>
<li><p>过程</p>
<ul>
<li>1.外植体脱分化形成愈伤组织</li>
<li><p>2.愈伤组织分散开形成细胞悬液</p>
<ul>
<li>纤维素酶和果胶酶</li>
<li>不分化</li>
</ul>
</li>
<li><p>3.发酵提取细胞产物</p>
</li>
</ul>
</li>
</ul>
</li>
</ul>
<h2 id="动物细胞工程"><a href="#动物细胞工程" class="headerlink" title="动物细胞工程"></a>动物细胞工程</h2><h3 id="动物细胞培养和核移植技术"><a href="#动物细胞培养和核移植技术" class="headerlink" title="动物细胞培养和核移植技术"></a>动物细胞培养和核移植技术</h3><ul>
<li><p>动物细胞培养</p>
<ul>
<li><p>原理</p>
<ul>
<li>有丝分裂</li>
</ul>
</li>
<li><p>概念</p>
<ul>
<li><p>从动物机体中取出相关的组织，将它们分散成单个细胞，然后，放在适宜的培养基中，让这些细胞生长和增殖</p>
<ul>
<li>取出的一般是胚胎或幼龄动物的器官组织</li>
</ul>
</li>
</ul>
</li>
<li><p>过程（见书P45图2-16）</p>
</li>
<li><p>条件</p>
<ul>
<li><p>无菌、无毒</p>
<ul>
<li>培养液和培养用具无菌处理</li>
<li>添加一定量的抗生素</li>
<li>定期更换培养液</li>
</ul>
</li>
<li><p>营养</p>
<ul>
<li>糖、氨基酸、促生长因子、无机盐、微量元素等</li>
<li>血清、血浆</li>
</ul>
</li>
<li><p>温度和pH</p>
<ul>
<li>哺乳动物多以36.5±0.5℃为宜</li>
<li>多数细胞适宜pH为7.2~7.4</li>
</ul>
</li>
<li><p>气体环境</p>
<ul>
<li>二氧化碳主要作用是维持培养液的pH</li>
<li>采用培养皿或者松盖培养瓶，置于含95%空气加5%二氧化碳的混合气体的培养箱中</li>
</ul>
</li>
</ul>
</li>
<li><p>应用</p>
</li>
</ul>
</li>
<li><p>动物体细胞核移植技术和克隆动物</p>
<ul>
<li><p>概念</p>
<ul>
<li>动物核移植是将动物的一个细胞的细胞核，移入一个已经去掉细胞核的卵母细胞中，使其重组并发育成一个新的胚胎，这个新的胚胎最终发育为动物个体。用核移植的方法得到的动物称为克隆动物。</li>
</ul>
</li>
<li><p>哺乳动物核移植分类</p>
<ul>
<li>胚胎细胞核移植</li>
<li>体细胞核移植</li>
</ul>
</li>
<li><p>体细胞核移植的过程（见书P48图2-21）</p>
<ul>
<li><p>去核</p>
<ul>
<li><p>显微操作去核法（普遍使用）</p>
<ul>
<li>微型吸管一并吸出细胞核与第一极体</li>
</ul>
</li>
<li><p>梯度离心、紫外光段时间招商、化学物质处理</p>
<ul>
<li>不刺破透明带或卵母细胞膜</li>
</ul>
</li>
</ul>
</li>
<li><p>选MⅡ期卵母细胞</p>
<ul>
<li>体积大，便于操作</li>
<li>含有促进细胞核全能性表达的物质和营养条件</li>
</ul>
</li>
<li><p>注入时直接将整个供体细胞注入去核卵母细胞</p>
</li>
<li><p>先促进两细胞融合</p>
<ul>
<li>电刺激</li>
</ul>
</li>
<li><p>再激活受体细胞</p>
<ul>
<li>物理方法：电脉冲</li>
<li>化学方法：钙离子载体、乙醇、蛋白酶合成抑制剂</li>
</ul>
</li>
</ul>
</li>
<li><p>应用前景</p>
<ul>
<li>促进优良畜种繁育</li>
<li>保护濒危物种</li>
<li>生产珍贵医用蛋白</li>
<li>组织器官移植</li>
</ul>
</li>
<li><p>存在的问题</p>
<ul>
<li>目前成功率低</li>
<li>克隆动物存在健康问题</li>
</ul>
</li>
</ul>
</li>
</ul>
<h3 id="动物细胞融合与单克隆抗体"><a href="#动物细胞融合与单克隆抗体" class="headerlink" title="动物细胞融合与单克隆抗体"></a>动物细胞融合与单克隆抗体</h3><ul>
<li><p>动物细胞融合</p>
<ul>
<li>概念：又称细胞杂交，是指两个或多个动物细胞结合形成一个细胞的过程</li>
<li>得到的称为“杂交细胞”</li>
<li><p>方法</p>
<ul>
<li>聚乙二醇PEG</li>
<li>灭活的病毒</li>
<li>电激</li>
</ul>
</li>
<li><p>意义</p>
</li>
</ul>
</li>
<li><p>单克隆抗体</p>
<ul>
<li><p>单克隆抗体的制备（书P53）</p>
<ul>
<li><p>选择两次</p>
<ul>
<li>先用特定的选择性培养基筛选出杂交瘤细胞</li>
<li>再克隆化培养、抗体检测</li>
</ul>
</li>
<li><p>最后</p>
<ul>
<li>在体外条件下大规模培养</li>
<li>注射到小鼠腹腔内增殖</li>
</ul>
</li>
</ul>
</li>
<li><p>单克隆抗体的应用</p>
<ul>
<li><p>作为诊断试剂</p>
<ul>
<li>抗原-抗体杂交</li>
<li>单克隆抗体纯度高、特异性强</li>
<li>诊断准确、高效、简易、快速</li>
</ul>
</li>
<li><p>用于治疗疾病和运载药物</p>
<ul>
<li><p>生物导弹</p>
<ul>
<li>疗效高、毒副作用小</li>
</ul>
</li>
</ul>
</li>
</ul>
</li>
</ul>
</li>
</ul>

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          <div class="post-toc motion-element"><ol class="nav"><li class="nav-item nav-level-1"><a class="nav-link" href="#细胞工程"><span class="nav-number">1.</span> <span class="nav-text">细胞工程</span></a><ol class="nav-child"><li class="nav-item nav-level-2"><a class="nav-link" href="#前言"><span class="nav-number">1.1.</span> <span class="nav-text">前言</span></a><ol class="nav-child"><li class="nav-item nav-level-3"><a class="nav-link" href="#原理和方法"><span class="nav-number">1.1.1.</span> <span class="nav-text">原理和方法</span></a></li><li class="nav-item nav-level-3"><a class="nav-link" href="#操作水平"><span class="nav-number">1.1.2.</span> <span class="nav-text">操作水平</span></a></li><li class="nav-item nav-level-3"><a class="nav-link" href="#目的"><span class="nav-number">1.1.3.</span> <span class="nav-text">目的</span></a></li><li class="nav-item nav-level-3"><a class="nav-link" href="#两大领域"><span class="nav-number">1.1.4.</span> <span class="nav-text">两大领域</span></a></li></ol></li><li class="nav-item nav-level-2"><a class="nav-link" href="#植物细胞工程"><span class="nav-number">1.2.</span> <span class="nav-text">植物细胞工程</span></a><ol class="nav-child"><li class="nav-item nav-level-3"><a class="nav-link" href="#理论基础"><span class="nav-number">1.2.1.</span> <span class="nav-text">理论基础</span></a></li><li class="nav-item nav-level-3"><a class="nav-link" href="#相关概念"><span class="nav-number">1.2.2.</span> <span class="nav-text">相关概念</span></a></li><li class="nav-item nav-level-3"><a class="nav-link" href="#基本技术"><span class="nav-number">1.2.3.</span> <span class="nav-text">基本技术</span></a></li><li class="nav-item nav-level-3"><a class="nav-link" href="#实际应用"><span class="nav-number">1.2.4.</span> <span class="nav-text">实际应用</span></a></li></ol></li><li class="nav-item nav-level-2"><a class="nav-link" href="#动物细胞工程"><span class="nav-number">1.3.</span> <span class="nav-text">动物细胞工程</span></a><ol class="nav-child"><li class="nav-item nav-level-3"><a class="nav-link" href="#动物细胞培养和核移植技术"><span class="nav-number">1.3.1.</span> <span class="nav-text">动物细胞培养和核移植技术</span></a></li><li class="nav-item nav-level-3"><a class="nav-link" href="#动物细胞融合与单克隆抗体"><span class="nav-number">1.3.2.</span> <span class="nav-text">动物细胞融合与单克隆抗体</span></a></li></ol></li></ol></li></ol></div>
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